Synaptic mechanisms of inspiratory off-switching evoked by pontine pneumotaxic stimulation in cats

To elucidate synaptic mechanisms and the involvement of N-methyl-D-aspartate (NMDA) receptors in inspiratory off-switching (IOS) evoked by the stimulation of the nucleus parabrachialis medialis (NPBM), excitatory and inhibitory postsynaptic potentials (EPSPs and IPSPs) were recorded from bulbar augmenting inspiratory (aug-I) and postinspiratory (PI) neurons in vagotomized cats. Stimulation of NPBM produced either transient inhibition or premature termination of inspiration (reversible or irreversible IOS), depending on the stimulus intensity. Each neuron displayed four-phasic postsynaptic responses during the reversible IOS, i.e. Phase 1 EPSPs, Phase 2 IPSPs, Phase 3 EPSPs and Phase 4 IPSPs in aug-I neurons, and Phase 1 plus 2 EPSPs, Phase 3 IPSPs and Phase 4 EPSPs in PI neurons. During the irreversible IOS, Phase 4 responses were replaced by sustained hyperpolarization in aug-I neurons and decrementing depolarization in PI neurons. Blockade of NMDA receptors by dizocilpine (0.3 mg kg(-1) i.v.) selectively increased Phase 4 potentials in both types of neurons and decreased the thresholds for evoking the irreversible IOS. The NPBM-induced responses had a pattern and time-course similar to those induced by vagal stimulation. The present results suggest that pneumotaxic and vagal inputs converge on the common IOS circuit, and the effectiveness of both inputs is modulated by NMDA receptors.     

Synaptic interactions between respiratory neurons during inspiratory on-switching evoked by vagal stimulation in decerebrate cats

To elucidate neuronal mechanisms underlying phase-switching from expiration to inspiration, or inspiratory on-switching (IonS), postsynaptic potentials (PSPs) of bulbar respiratory neurons together with phrenic nerve discharges were recorded during IonS evoked by vagal stimulation in decerebrate and vagotomized cats. A single shock stimulation of the vagus nerve applied at late-expiration developed an inspiratory discharge in the phrenic neurogram after a latency of 79+/-11 ms (n = 11). Preceding this evoked inspiratory discharge, a triphasic response was induced, consisting of an early silence (phase 1 silence), a transient burst discharge (phase 2 discharge) and a late pause (phase 3 pause). During phase 1 silence, IPSPs occurred in augmenting inspiratory (aug-I) and expiratory (E2) neurons, and EPSPs in postinspiratory (PI) neurons. During phase 2 discharge, EPSPs arose in aug-I neurons and IPSPs in PI and E2 neurons. These initial biphasic PSPs were comparable with those during inspiratory off-switching evoked by the same stimulation applied at late-inspiration. In both on- and off-switching, phase-transition in respiratory neuronal activities started to arise concomitantly with the phrenic phase 3 pause. These results suggest that vagal inputs initially produce a non-specific, biphasic response in bulbar respiratory neurons, which consecutively activates a more specific process connected to IonS.     

Peripheral chemoreceptor inputs to medullary inspiratory and postinspiratory neurons of cats

The effect of peripheral chemoreceptor activation on inspiratory and postinspiratory medullary neurons was investigated using intracellular recording techniques. Peripheral chemoreceptors were activated by injecting CO₂ saturated 1 N bicarbonate solution into the lingual artery or by electrically stimulating the carotid sinus nerve. Injections of 20–300 l bicarbonate solution evoked changes in respiratory frequency and in peak phrenic nerve discharge. The membrane potential of inspiratory alpha neurons, whether bulbospinal or not and independent of their anatomic location, was decreased during inspiration. A sequence of compound excitatory and inhibitory effects were observed when the stimulus was given during the postinspiratory and expiratory phases of the respiratory cycle. Inspiratory beta- and late-inspiratory neurons, however, were inhibited by peripheral chemoreceptor activation. Postinspiratory neurons were strongly activated during postinspiration. Neither class of respiratory neurons were shown to receive direct synaptic inputs from the peripheral chemoreceptors as tested by electrical stimulation of the carotid sinus nerve and signal averaging of the respiratory neuron membrane potential. The experiments revealed differential influences of afferent chemoreceptor activity on various components of the respiratory network. We conclude that chemoreceptor afferents activate non-respiratory modulated medullary neurons which, in turn, activate or inhibit various neurons of the medullary respiratory control network. The responses of each type of respiratory neuron to chemoreceptors afferents may then be considered in the context of this direct interaction as well as the network interactions of the various cells.     

Modulation of synaptic transmission in hippocampal CA1 neurons by a novel neurotoxin (beta-pompilidotoxin) derived from wasp venom.

We examined the effects of beta-pompilidotoxin (beta-PMTX), a neurotoxin derived from wasp venom, on synaptic transmission in the mammalian central nervous system (CNS). Using hippocampal slice preparations of rodents, we made both extracellular and intracellular recordings from the CA1 pyramidal neurons in response to stimulation of the Schaffer collateral/commissural fibers. Application of 5-10 microM beta-PMTX enhanced excitatory postsynaptic potentials (EPSPs) but suppressed the fast component of the inhibitory postsynaptic potentials (IPSPs). In the presence of 10 microM bicuculline, beta-PMTX potentiated EPSPs that were composed of both non-NMDA and NMDA receptor-mediated potentials. Potentiation of EPSPs was originated by repetitive firings of the presynaptic axons, causing summation of EPSPs. In the presence of 10 microM CNQX and 50 microM APV, beta-PMTX suppressed GABA(A) receptor-mediated fast IPSPs but retained GABA(B) receptor-mediated slow IPSPs. Our results suggest that beta-PMTX facilitates excitatory synaptic transmission by a presynaptic mechanism and that it causes overexcitation followed by block of the activity of some population of interneurons which regulate the activity of GABA(A) receptors.     

Excitation and inhibition of medullary inspiratory neurons by two types of burst inspiratory neurons in the cat

In Nembutal-anesthetized and artificially ventilated cats, we studied the connectivity of burst inspiratory (I) neurons in the B?tzinger complex and the ventral respiratory group (VRG) with spike-triggered averaging methods. Burst I neurons exhibited tonic (I-TON) or decrementing (I-DEC) firing patterns. Spikes of I-TON neurons induced monosynaptic EPSPs in intracellularly recorded I neurons of both the VRG and the dorsal respiratory group (DRG). Spikes of I-DEC neurons induced monosynaptic inhibitory postsynaptic potentials (IPSPs) in both VRG and DRG I neurons.     

Differential inhibitory control of semicircular canal nerve afferent-evoked inputs in second-order vestibular neurons by glycinergic and GABAergic circuits

Labyrinthine nerve-evoked monosynaptic excitatory postsynaptic potentials (EPSPs) in second-order vestibular neurons (2°VN) sum with disynaptic inhibitory postsynaptic potentials (IPSPs) that originate from the thickest afferent fibers of the same nerve branch and are mediated by neurons in the ipsilateral vestibular nucleus. Pharmacological properties of the inhibition and the interaction with the afferent excitation were studied by recording monosynaptic responses of phasic and tonic 2°VN in an isolated frog brain after electrical stimulation of individual semicircular canal nerves. Specific transmitter antagonists revealed glycine and GABA_A receptor-mediated IPSPs with a disynaptic onset only in phasic but not in tonic 2°VN. Compared with GABAergic IPSPs, glycinergic responses in phasic 2°VN have larger amplitudes and a longer duration and reduce early and late components of the afferent nerve-evoked subthreshold activation and spike discharge. The difference in profile of the disynaptic glycinergic and GABAergic inhibition is compatible with the larger number of glycinergic as opposed to GABAergic terminal-like structures on 2°VN. The increase in monosynaptic excitation after a block of the disynaptic inhibition in phasic 2°VN is in part mediated by a N-methyl-D-aspartate receptor-activated component. Although inhibitory inputs were superimposed on monosynaptic EPSPs in tonic 2°VN as well, the much longer latency of these IPSPs excludes a control by short-latency inhibitory feed-forward side-loops as observed in phasic 2°VN. The differential synaptic organization of the inhibitory control of labyrinthine afferent signals in phasic and tonic 2°VN is consistent with the different intrinsic signal processing modes of the two neuronal types and suggests a co-adaptation of intrinsic membrane properties and emerging network properties.     

Converse motor output of inspiratory bulbospinal premotoneurones during vomiting

Intracellular recordings of the activities of 10 inspiratory bulbospinal neurones of the medulla were performed in decerebrate cats. Fictive vomiting was induced by repetitive stimulation of the supra-diaphragmatic vagus nerves and was defined by series of synchronous large bursts in both the phrenic (inspiratory) and abdominal (expiratory) nerves. During these synchronous bursts the inspiratory bulbospinal neurones of both the dorsal and ventral respiratory groups were strongly hyperpolarized by chloride-dependent inhibitory postsynaptic potentials (IPSPs). We concluded that during vomiting the central pattern generator is inhibited, and that another pattern generator drives the spinal respiratory motoneurones.     

Pneumotaxic mechanisms in the non-human primate: effect of the N-methyl-D-aspartate (NMDA) antagonist ketamine

We tested the possible involvement of N-methyl-D-aspartate (NMDA) receptors in the central inspiratory-termination mechanism in non-human primates. Inspiratory bursts were recorded from the phrenic nerve in Macaca fascicularis monkeys paralyzed and ventilated by means of a servoventilator driven by the inspiratory discharge of the phrenic nerve. The central inspiratory termination mechanism was tested by withholding lung inflation. This transiently suppressed the vagal feedback from the lungs which produces inspiratory off-switching independent from the central mechanism. Under anaesthesia with ketamine, a potent NMDA antagonist, non inflation increased inspiratory time to 4s (1s with lungs inflated) whereas no such effect was observed during halothane anaesthesia. We conclude that the termination of inspiration in primates is controlled via central mechanisms in which NMDA receptors are involved.     

Activity of dorsal respiratory group inspiratory neurons during laryngeal-induced fictive coughing and swallowing in decerebrate cats

Membrane potential changes and/or discharges from 36 inspiratory neurons were recorded intracellularly in the dorsal respiratory group (DRG; i.e., the ventrolateral subdivision of the nucleus tractus solitarii) in decerebrate, paralyzed, and ventilated cats. Electrical activities were recorded from both somata (n=10) and axons (n=26). Activities during quiet breathing were compared with those observed during fictive coughing and swallowing evoked by repetitive electrical stimulation of afferent fibers of the superior laryngeal nerve (SLN). These nonrespiratory behaviors were evident in paralyzed animals as characteristic discharge patterns of the phrenic, abdominal, and hypoglossal nerves. Twenty-six neurons exhibiting antidromic action potentials in response to electrical stimuli applied to the cervical (C3–5) spinal cord were classified as inspiratory bulbospinal neurons (IBSNs). These neurons were considered as premotoneurons. The remaining 10 inspiratory neurons (INAA) were not antidromically activated by electrical stimuli applied to either cervical spinal cord or ipsilateral cervical vagus. These neurons are thought to be propriobulbar neurons. We recorded the activity of 31 DRG inspiratory neurons (24 IBSNs and 7 I-NAA) during coughing. All but one (a late-recruited IBSN) discharged a burst of action potentials during the coughing-related phrenic nerve activity. Typically, ramp-like membrane depolarization trajectories and discharge frequencies during coughing were similar to those observed during inspiration. We recorded the activity of 33 DRG inspiratory neurons (23 IBSNs and 10 I-NAA) during swallowing. Most (28/33) neurons were briefly activated, i.e., discharged a burst of action potentials during swallowing, but peak discharge frequency decreased compared with that measured during inspiration. The membrane potentials of nine somata exhibited a brief bell-shaped depolarization during swallowing, the amplitude of which was similar to that observed during inspiration. These results suggest that some inspiratory premotoneurons and propriobulbar neurons of the DRG might be involved in nonrespiratory motor activities, even if clearly antagonistic to breathing (e.g., swallowing). We postulate the existence in the medulla oblongata of adult mammals of neurons exhibiting a functional flexibility.     

Dynamic modulation of excitation and inhibition during stimulation at gamma and beta frequencies in the CA1 hippocampal region

Fast oscillations at gamma and beta frequency are relevant to cognition. During this activity, excitatory and inhibitory postsynaptic potentials (EPSPs and IPSPs) are generated rhythmically and synchronously and are thought to play an essential role in pacing the oscillations. The dynamic changes occurring to excitatory and inhibitory synaptic events during repetitive activation of synapses are therefore relevant to fast oscillations. To cast light on this issue in the CA1 region of the hippocampal slice, we used a train of stimuli, to the pyramidal layer, comprising 1 s at 40 Hz followed by 2--3 s at 10 Hz, to mimic the frequency pattern observed during fast oscillations. Whole cell current-clamp recordings from CA1 pyramidal neurons revealed that individual stimuli at 40 Hz produced EPSPs riding on a slow biphasic hyperpolarizing-depolarizing waveform. EPSP amplitude initially increased; it then decreased concomitantly with the slow depolarization and with a large reduction in membrane resistance. During the subsequent 10-Hz train: the cells repolarized, EPSP amplitude and duration increased to above control, and no IPSPs were detected. In the presence of GABA(A) receptor antagonists, the slow depolarization was blocked, and EPSPs of constant amplitude were generated by 10-Hz stimuli. Altering pyramidal cell membrane potential affected the time course of the slow depolarization, with the peak being reached earlier at more negative potentials. Glial recordings revealed that the trains were associated with extracellular potassium accumulation, but the time course of this event was slower than the neuronal depolarization. Numerical simulations showed that intracellular chloride accumulation (due to massive GABAergic activation) can account for these observations. We conclude that synchronous activation of inhibitory synapses at gamma frequency causes a rapid chloride accumulation in pyramidal neurons, decreasing the efficacy of inhibitory potentials. The resulting transient disinhibition of the local network leads to a short-lasting facilitation of polysynaptic EPSPs. These results set constraints on the role that synchronous, rhythmic IPSPs may play in pacing oscillations at gamma frequency in the CA1 hippocampal region.     

Electrophysiological and pharmacological analysis of synaptic inputs to pulmonary rapidly adapting receptor relay neurons in the rat

The information from pulmonary rapidly adapting stretch receptors (RARs) to the central nervous system (CNS) is relayed in the nucleus tractus solitarii (NTS). The second-order neurons in the NTS referred to as RAR cells have recently been shown to receive rhythmic inputs from the central respiratory system in addition to the main inputs from RAR afferents. The present study analyzed these synaptic inputs by intracellular recordings from RAR cells, and by extracellular recordings combined with local applications of neuroactive drugs to RAR cells, in Nembutal-anesthetized, paralyzed, and artificially ventilated rats. The intracellular analysis identified both excitatory postsynaptic potentials (EPSPs) elicited presumably by RAR afferents and inhibitory postsynaptic potentials (IPSPs) synchronous with central inspiratory activity. This inhibitory input, called I suppression, was the origin of respiratory modulation of RAR cell firing, and its time course suggested that some unidentified inspiratory neurons with an augmenting firing pattern were the source of the inhibition. The pharmacological analysis suggested the types of neurotransmitters used in these synaptic events. First, glutamate was shown to be the primary neurotransmitter at the synapse between RAR afferents and RAR cells. Iontophoretic applications of the non-NMDA glutamate antagonist, CNQX, abolished RAR cell firing almost completely in response to lung inflation and deflation and to electrical stimulation of the vagus nerve. Second, glycinergic inputs which inhibited RAR cells in the inspiratory phase were revealed by applications of the glycine antagonist, strychnine. That is, the I suppression was greatly diminished by applications of strychnine. Third, although applications of the GABA_A receptor antagonist, bicuculline, had little effect on I suppression, bicuculline markedly increased the baseline firing of RAR cells. These results imply that the information path from RARs to the CNS is regulated at the level of RAR cells by phasically-acting glycinergic inhibition in the inspiratory phase and tonically-acting GABAergic inhibition; the results also provide new insights into the neuronal mechanisms of RAR-induced reflexes. Received: 3 February 1999 / Accepted: 12 May 1999     

Inhibitory component of the resistance reflex in the locomotor network of the crayfish

The aim of this study was to investigate the inhibitory components of a resistance reflex in the walking system of the crayfish. This study was performed using an in vitro preparation of several thoracic ganglia including motor nerves and the proprioceptor that codes movements of the second joint (coxo-basipodite chordotonal organ-CBCO). Sinusoidal movements were imposed on the CBCO, and intracellular responses were recorded from levator (Lev) and depressor (Dep) motoneurons (MNs). We found that in MNs that oppose the imposed movements (e.g., the Lev MNs during the imposed downward movement), the response consists in a depolarization resulting from the summation of excitatory postsynaptic potentials (EPSPs). A movement in the opposite direction resulted in hyperpolarization during which inhibitory postsynaptic potentials (IPSPs) summated. The inhibitory pathway to each MN is oligosynaptic (i.e., composed of a small number of neurons in series) and involves spiking interneurons because it was blocked in the presence of a high-divalent cation solution. The IPSPs were mediated by a chloride conductance because their amplitude was sensitive to the chloride concentration of the bathing solution and because they were blocked by the chloride channel blocker, picrotoxin. Resistance reflex IPSPs related to single CBCO neurons could be identified. These unitary IPSPs were blocked in the presence of 3-mercapto-propionic acid, an inhibitor of gamma-amino-butyric acid (GABA) synthesis, indicating that they are mediated by GABA. In addition to this GABAergic pathway, electrical stimulation of the CBCO sensory nerve induced compound IPSPs that were blocked by glutamate pyruvate transaminase (GPT), indicating the presence of glutamatergic inhibitory pathways. These glutamatergic interneurons do not appear to be involved in the resistance reflex, however, as GPT did not block the unitary IPSPs. Functionally, the resistance reflex is mainly supported by movement-coding CBCO sensory neurons. We demonstrate that such movement-coding CBCO neurons produce both monosynaptic EPSPs in the MNs opposing imposed movements and oligosynaptic IPSPs in the antagonistic motoneurons. These results highlight the similarities between the inhibitory pathways in resistance reflex of the crayfish and in the stretch reflex of vertebrates mediated by Ia inhibitory interneurons.     

Differential sensitivity of excitatory and inhibitory synaptic transmission to modulation by nitric oxide in rat nucleus tractus solitarii

The nucleus tractus solitarii (NTS) is a key central link in control of multiple homeostatic reflexes. A number of studies have demonstrated that exogenous and endogenous nitric oxide (NO) within NTS regulates visceral function, but further understanding of the role of NO in the NTS is hampered by the lack of information about its intracellular actions. We studied effects of NO in acute rat brainstem slices. Aqueous NO solution (NO(aq)) potentiated electrically evoked excitatory and inhibitory postsynaptic potentials (EPSPs and IPSPs, respectively) in different neuronal subpopulations and, in some neurones, caused a depolarization. Similar effects were observed using the NO donor diethylamine NONOate (DEA/NO). The threshold NO concentration as determined using an NO electrochemical sensor was estimated as approximately 0.4 nm (EC(50) approximately 0.9 nm) for potentiating glutamatergic EPSPs but approximately 3 nm for monosynaptic GABAergic IPSPs. Bath application of the soluble guanylate cyclase (sGC) inhibitor 1H-[1,2,4]oxadiazolo[4,3-a]quinoxalin-1-one (ODQ) abolished NO(aq)- and DEA/NO-induced potentiation of evoked EPSPs, IPSPs and depolarization. All NO actions were mimicked by the non-NO-dependent guanylate cyclase activator Bay 41-2272. The effects of NO on EPSPs and IPSPs persisted in cells where postsynaptic sGC was blocked by ODQ and therefore were presynaptic, owing to a direct modulation of transmitter release combined with depolarization of presynaptic neurones. Therefore, while lower concentrations of NO may be important for fine tuning of glutamatergic transmission, higher concentrations are required to directly engage GABAergic inhibition. This differential sensitivity of excitatory and inhibitory connections to NO may be important for determining the specificity of the effects of this freely diffusible gaseous messenger.     

Synaptic responses of guinea pig cingulate cortical neurons in vitro.

1. Intracellular recordings were made from layer V/VI neurons of the guinea pig anterior cingulate cortex to investigate postsynaptic potentials (PSPs) evoked by electrical stimulation of the subcortical white matter (forceps minor). 2. Four distinct types of PSPs were recorded (at the resting potential) under normal physiological conditions; 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX)-sensitive excitatory postsynaptic potentials (EPSPs) were followed by bicuculline- or picrotoxin-sensitive depolarizing or hyperpolarizing inhibitory postsynaptic potentials (IPSPs), which were further followed by phaclofen-sensitive, long-lasting hyperpolarizing postsynaptic potentials (LPSPs). The average times-to-peak for the EPSP, depolarizing and hyperpolarizing IPSPs, and LPSP were 10, 22, 28, and 146 ms, respectively. 3. In the presence of CNQX and bicuculline, high-intensity electrical stimulation elicited a longer lasting EPSP with a time-to-peak of 21 ms. The amplitude and duration of the EPSP decreased with membrane hyperpolarization and increased with membrane depolarization. The EPSP was reversibly abolished by D,L-2-amino-5-phosphonovaleric acid (D,L-APV). 4. The bicuculline- or picrotoxin-sensitive depolarizing and hyperpolarizing IPSPs and the phaclofen-sensitive LPSP were markedly suppressed by CNQX, suggesting that glutamate (Glu) and/or aspartate nerve terminals project to GABAergic interneurons, and that the GABAergic interneurons are activated mainly by non-N-methyl-D-aspartate (non-NMDA) receptors. 5. In the presence of picrotoxin, the average reversal potential for the compound EPSP was 0 mV, which was similar to that (-6 mV) for the Glu-induced depolarization. In a solution containing D,L-APV at low concentrations, the average reversal potentials for the depolarizing and hyperpolarizing IPSPs and for the early and late components of the gamma-aminobutyric acid (GABA)-induced responses were -62, -72, -70, and -61 mV, respectively. Thus the value for the depolarizing IPSP was similar to that for the late response to GABA, whereas the value for the hyperpolarizing IPSP was almost the same as that for the early response to GABA. The average reversal potential of -90 mV for the LPSP was similar to -93 mV for the baclofen-induced hyperpolarization and to -94 mV for the spike afterhyperpolarization. 6. Application of phaclofen decreased the interspike interval of the spontaneous firing and reversed the increase in the interspike interval after subcortical stimulation. This result indicates that, even in a slice preparation, the anterior cingulate neurons are under tonic inhibitory control exerted by spontaneously active GABAergic interneurons.(ABSTRACT TRUNCATED AT 400 WORDS)     

Layer-specific pyramidal cell oscillations evoked by tetanic stimulation in the rat hippocampal area CA1 in vitro and in vivo

Tetanic stimulation of axons terminating in the CA1 region of the hippocampus induces oscillations in the gamma-to-beta frequency band (13–100 Hz) and can induce long-term potentiation (LTP). The rapid pyramidal cell discharge is driven by a mainly GABA_A-receptor-mediated slow depolarization and entrained mainly through ephaptic interactions. This study tests whether cellular compartmentalization can explain how cells, despite severely reduced input resistance, can still fire briskly and have IPSPs superimposed on the slow GABAergic depolarization, and whether this behaviour occurs in vivo . Oscillations induced in CA1 in vitro by tetanic stimulation of the stratum radiatum or oriens were analysed using intracellular and multichannel field potentials along the cell axis. Layer-specific effects of focal application of bicuculline indicate that the GABAergic depolarization is concentrated on tetanized dendrites. Current-source density analysis and characteristics of partial spikes indicate that early action potentials are initiated in the proximal nontetanized dendrite but cannot invade the tetanized dendrite, where recurrent EPSPs and evoked IPSPs were largely suppressed. As the oscillation progresses, IPSPs recover and slow the neuronal firing to β frequencies, with a small subpopulation of neurons continuing to fire at γ frequency. Carbonic anhydrase dependence, threshold intensity, frequency, field strength and spike initiation/propagation of tetanus-evoked oscillations in urethane-anaesthetized rats, validate our observations in vitro , and show that these mechanisms operate in healthy tissue. However, the disrupted electrophysiology of the tetanized dendrites will disable normal information processing, has implications for LTP induction and is likely to play a role in pathological synchronization as found during epileptic discharges.     

Synaptic responses of neurons controlling the parotid and von Ebner salivary glands in rats to stimulation of the solitary nucleus and tract

Salivary secretion results from reflex stimulation of autonomic neurons via afferent sensory information relayed to neurons in the rostral nucleus of the solitary tract (rNST), which synapse with autonomic neurons of the salivatory nuclei. We investigated the synaptic properties of the afferent sensory connection to neurons in the inferior salivatory nucleus (ISN) controlling the parotid and von Ebner salivary glands. Mean synaptic latency recorded from parotid gland neurons was significantly shorter than von Ebner gland neurons. Superfusion of GABA and glycine resulted in a concentration-dependent membrane hyperpolarization. Use of glutamate receptor antagonists indicated that both AMPA and N-methyl-D-aspartate (NMDA) receptors are involved in the evoked excitatory postsynaptic potentials (EPSPs). Inhibitory postsynaptic potential (IPSP) amplitude increased with higher intensity ST stimulation. Addition of the glycine antagonist strychnine did not affect the amplitude of the IPSPs significantly. The GABA(A) receptor antagonist, bicuculline (BMI) or mixture of strychnine and BMI abolished the IPSPs in all neurons. IPSP latency was longer than EPSP latency, suggesting that more than one synapse is involved in the inhibitory pathway. Results show that ISN neurons receive both excitatory and inhibitory afferent input mediated by glutamate and GABA respectively. The ISN neuron response to glycine probably derives from descending connections. Difference in the synaptic characteristics of ISN neurons controlling the parotid and von Ebner glands may relate to the different function of these two glands.     

Kindling-induced long-lasting changes in synaptic transmission in the basolateral amygdala.

1. Intracellular current-clamp recordings were obtained from neurons of the basolateral amygdala (BLA) in an in vitro slice preparation from control and kindled animals. Postsynaptic potentials, elicited by stimulation of the stria terminalis (ST) or lateral amygdaloid nucleus (LA), were used to investigate the role of excitatory and inhibitory amino acid transmission in kindling-induced epileptiform activity. The contributions of glutamatergic and GABAergic receptor subtypes were analyzed by use of the non-N-methyl-D-aspartate (non-NMDA) antagonist 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX), the NMDA antagonist DL-2-amino-5-phosphonovaleric acid (APV), and the GABAA antagonist bicuculline methiodide (BMI). 2. The synaptic waveform evoked in control neurons consisted of an excitatory postsynaptic potential (EPSP), a fast inhibitory postsynaptic potential (f-IPSP), and a slow inhibitory postsynaptic potential (s-IPSP). Stimulation of the ST or LA pathways evoked a burst-firing response in BLA neurons contralateral from the site of stimulation of kindled animals. 3. APV (50 microM) reduced, but CNQX (10 microM) completely blocked, the burst-firing response in BLA neurons from kindled animals and bicuculline-induced bursting in control neurons. 4. Kindling significantly increased the amplitude of both the slow NMDA- and the fast non-NMDA-receptor-mediated components of synaptic transmission (s- and f-EPSPs, respectively). Furthermore, the stimulus intensities required to evoke EPSPs just subthreshold for action potential generation were significantly lower in slices from kindled animals. 5. In kindled neurons no significant change was observed in the membrane input resistance and resting membrane potential or in the number of action potentials elicited in response to depolarizating current injection. 6. Kindling resulted in a pathway-specific loss of ST- and LA-evoked feedforward GABAergic synaptic transmission and of spontaneous IPSPs. In the same BLA neurons, direct GABAergic inhibition via stimulation of the LA was not affected by kindling. 7. The enhanced glutamatergic transmission was not due to disinhibition, because, in the presence of BMI (and CNQX to prevent BMI-induced bursting), the s-EPSP amplitude was still greater in kindled than in control neurons. 8. These results provide evidence that the epileptiform activity observed in BLA neurons after kindling results from an increase in excitatory NMDA- and non-NMDA-receptor-mediated glutamatergic transmission and a decrease in inhibitory gamma-aminobutyric acid (GABA)-receptor-mediated transmission; the enhanced excitatory transmission cannot be accounted for by reduced inhibition.(ABSTRACT TRUNCATED AT 400 WORDS)     

Excitatory postsynaptic potentials in rat neocortical neurons in vitro. III. Effects of a quinoxalinedione non-NMDA receptor antagonist

1. Intracellular microelectrodes were used to obtain recordings from neurons in layer II/III of rat frontal cortex. A bipolar electrode positioned in layer IV of the neocortex was used to evoke postsynaptic potentials. Graded series of stimulation were employed to selectively activate different classes of postsynaptic responses. The sensitivity of postsynaptic potentials and iontophoretically applied neurotransmitters to the non-N-methyl-D-asparate (NMDA) antagonist 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX) was examined. 2. As reported previously, low-intensity electrical stimulation of cortical layer IV evoked short-latency early excitatory postsynaptic potentials (eEPSPs) in layer II/III neurons. CNQX reversibly antagonized eEPSPs in a dose-dependent manner. Stimulation at intensities just subthreshold for activation of inhibitory postsynaptic potentials (IPSPs) produced long-latency (10 to 40-ms) EPSPs (late EPSPs or 1EPSPs). CNQX was effective in blocking 1EPSPs. 3. With the use of stimulus intensities at or just below threshold for evoking an action potential, complex synaptic potentials consisting of EPSP-IPSP sequences were observed. Both early, Cl(-)-dependent and late, K(+)-dependent IPSPs were reduced by CNQX. This effect was reversible on washing. This disinhibition could lead to enhanced excitability in the presence of CNQX. 4. Iontophoretic application of quisqualate produced a membrane depolarization with superimposed action potentials, whereas NMDA depolarized the membrane potential and evoked bursts of action potentials. At concentrations up to 5 microM, CNQX selectively antagonized quisqualate responses. NMDA responses were reduced by 10 microM CNQX. D-Serine (0.5-2 mM), an agonist at the glycine regulatory site on the NMDA receptor, reversed the CNQX depression of NMDA responses.(ABSTRACT TRUNCATED AT 250 WORDS)     

Synchronization of ventral-group, bulbospinal inspiratory neurons in the decerebrate rat

We examined the synaptic connections between ventral-group, bulbospinal inspiratory neurons in 27 vagotomized, paralyzed, ventilated, and decerebrated rats using cross-correlation and spike-triggered averaging of intracellular potentials. The neurons were recorded in the medulla about the level of the obex and identified by their inspiratory firing pattern and antidromic activation from the spinal cord at C7. Whole C5 phrenic nerve recordings were made using bipolar electrodes from the central cut ends of the nerve. Most (108/137, 79%) inspiratory neurons discharged only during inspiration but some (29/137, 21%) also discharged during early expiration. Their intracellular membrane potentials displayed a pattern of depolarization during inspiration, repolarization during early expiration, and hyperpolarization during late expiration. Intracellular chloride iontophoresis changed the inspiratory membrane potential trajectories from augmenting to decrementing in 11 of 19 neurons tested (58%), and demonstrated the presence of both early-decrementing and late-augmenting waves of inhibitory postsynaptic potentials during expiration in 11 of 19 neurons tested (58%). Cross-correlation histograms were computed between pairs of extracellularly recorded neurons to detect short time scale synchronizations indicative of synaptic connections (26 ipsilateral; 23 contralateral). While none of the cross-correlation histograms for contralateral pairs showed peaks, most (23, 88%) of those for ipsilateral pairs showed peaks (mean half-amplitude width ± SD = 1.3 ± 0.4 ms) at time zero suggestive of common activation. Some of the latter (6, 23%) showed troughs superimposed on the central peaks (mean half-amplitude width ± SD = 0.9 ± 0.2 ms) at short latencies (mean latency ± SD = 1.8 ± 1.9 ms) suggestive of inhibition; others (8, 31%) had asymmetrical central peaks and two had bilateral peaks suggesting more complex interconnections. Averages of intracellular membrane potentials of inspiratory neurons (n = 24), triggered by action potentials of a nearby extracellularly recorded inspiratory neuron, were computed to detect synchronized postsynaptic potentials. Over half (16, 67%) showed postsynaptic potentials (mean amplitude ± SD = 201 ± 176 μV; mean half-amplitude width ± SD = 2.3 ± 0.8 ms) confirming the cross-correlation findings of common excitation. We conclude that in decerebrated rats, ventral-group inspiratory neurons projecting to the C7 spinal segment share powerful, ipsilaterally distributed excitatory inputs which enhance their synchronous activity during inspiration. They also receive inhibition during inspiration and early-decrementing and late-augmenting inhibitory inputs during expiration. Received: 25 October 1996 / Accepted: 21 March 1997     

EPSPs in rat neocortical neurons in vitro. II. Involvement of N-methyl-D-aspartate receptors in the generation of EPSPs

1. Intracellular recordings were obtained from neurons in layer II/III of rat frontal cortex. Single-electrode current- and voltage-clamp techniques were employed to compare the sensitivity of excitatory postsynaptic potentials (EPSPs) and iontophoretically evoked responses to N-methyl-D-aspartate (NMDA) to the selective NMDA antagonist D-2-amino-5-phosphonovaleric acid (D-2-APV). The voltage dependence of the amplitudes of the EPSPs before and after pharmacologic changes in the neuron's current-voltage relationship was also examined. 2. NMDA depolarized the membrane potential, increased the neuron's apparent input resistance (RN), and evoked bursts of action potentials. The NMDA-induced membrane current (INMDA) gradually increased with depolarization from -80 to -40 mV. The relationship between INMDA and membrane potential displayed a region of negative slope conductance in the potential range between -70 and -40 mV which was sufficient to explain the apparent increase in RN and the burst discharges during the NMDA-induced depolarization. 3. Short-latency EPSPs (eEPSPs) were evoked by low-intensity electrical stimulation of cortical layer IV. Changes in the eEPSP waveform following membrane depolarization and hyperpolarization resembled those of NMDA-mediated responses. However, the eEPSP was insensitive to D-2-APV applied at concentrations (up to 20 microM) that blocked NMDA responses. 4. EPSPs with latencies between 10 and 40 ms [late EPSPs (lEPSPs)] were evoked by electrical stimulation using intensities just subthreshold to the activation of IPSPs. The amplitude of the lEPSP increased with hyperpolarization and decreased with depolarization. 5. The lidocaine derivative QX-314, injected intracellularly, suppressed sodium-dependent action potentials and depolarizing inward rectification. Simultaneously, the amplitude of the eEPSP significantly decreased with depolarization. Neither the amplitude of a long-latency EPSP nor the amplitude of inhibitory postsynaptic potentials (IPSPs) was significantly affected by QX-314. 6. Cesium ions (0.5-2.0 mM) added to the bathing solution reduced or blocked hyperpolarizing inward rectification. Under these conditions, the amplitude of the eEPSP increased with hyperpolarization. The amplitude of the lEPSP was unaltered or enhanced. 7. The lEPSP was reversibly blocked by D-2-APV (5-20 microM), although the voltage-dependence of its amplitude did not resemble the action of NMDA on neocortical neurons.(ABSTRACT TRUNCATED AT 400 WORDS)